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Biomedical subjects

B Das

Publications and source records attributed to B Das.

At least 163 records · Page 9Linked to original sources
PubMed1985

Purification and properties of aldose reductase and aldehyde reductase II from human erythrocyte.

Aldose reductase (EC 1.1.1.21) and aldehyde reductase II (L-hexonate dehydrogenase, EC 1.1.1.2) have been purified to homogeneity from human erythrocytes by using ion-exchange chromatography, chromatofocusing, affinity chromatography, and Sephadex gel filtration. Both enzymes are monomeric, Mr 32,500, by the criteria of the Sephadex gel filtration and polyacrylamide slab gel electrophoresis under denaturing conditions. The isoelectric pH's for aldose reductase and aldehyde reductase II were determined to be 5.47 and 5.06, respectively. Substrate specificity studies showed that aldose reductase, besides catalyzing the reduction of various aldehydes such as propionaldehyde, pyridine-3-aldehyde and glyceraldehyde, utilizes aldo-sugars such as glucose and galactose. Aldehyde reductase II, however, did not use aldo-sugars as substrate. Aldose reductase activity is expressed with either NADH or NADPH as cofactors, whereas aldehyde reductase II can utilize only NADPH. The pH optima for aldose reductase and aldehyde reductase II are 6.2 and 7.0, respectively. Both enzymes are susceptible to the inhibition by p-hydroxymercuribenzoate and N-ethylmaleimide. They are also inhibited to varying degrees by aldose reductase inhibitors such as sorbinil, alrestatin, quercetrin, tetramethylene glutaric acid, and sodium phenobarbital. The presence of 0.4 M lithium sulfate in the assay mixture is essential for the full expression of aldose reductase activity whereas it completely inhibits aldehyde reductase II. Amino acid compositions and immunological studies further show that erythrocyte aldose reductase is similar to human and bovine lens aldose reductase, and that aldehyde reductase II is similar to human liver and brain aldehyde reductase II.

Aldehyde Reductase↗
PubMed1985

Effect of slate dust on the rat erythrocyte membrane composition: in vitro and in vivo studies.

The biochemical composition of rat erythrocyte ghost membrane obtained by hypotonic lysis and slate-dust-induced lysis were compared in vitro. The phospholipids and glycosamine contents decreased in slate-dust-exposed erythrocyte membrane, whereas there were no changes in protein content. The in vitro and in vivo association of silica, leaching from slate dust, with the components of rat erythrocyte ghost membrane, has also been demonstrated. The interaction of silica with membrane constituents is proposed as a mechanism of action of slate-dust toxicity.

Amino Sugars↗
PubMed1985

Activated and unactivated forms of human erythrocyte aldose reductase.

Aldose reductase (alditol:NADP+ 1-oxidoreductase, EC 1.1.1.21) has been partially purified from human erythrocytes by DEAE-cellulose (DE-52) column chromatography. This enzyme is activated severalfold upon incubation with 10 microM each glucose 6-phosphate, NADPH, and glucose. The activation of the enzyme was confirmed by following the oxidation of NADPH as well as the formation of sorbitol with glucose as substrate. The activated form of aldose reductase exhibited monophasic kinetics with both glyceraldehyde and glucose (Km of glucose = 0.68 mM and Km of glyceraldehyde = 0.096 mM), whereas the native (unactivated) enzyme exhibited biphasic kinetics (Km of glucose = 9.0 and 0.9 mM and Km of glyceraldehyde = 1.1 and 0.14 mM). The unactivated enzyme was strongly inhibited by aldose reductase inhibitors such as sorbinil, alrestatin, and quercetrin, and by phosphorylated intermediates such as ADP, glycerate 3-phosphate, glycerate 1,3-bisphosphate, and glycerate 2,3-trisphosphate. The activated form of the enzyme was less susceptible to inhibition by aldose reductase inhibitors and phosphorylated intermediates.

Aldehyde Reductase↗
PubMed1985

Activation of aldose reductase from human tissues.

Human aorta, brain, and muscle aldose reductase, partially purified by DEAE-cellulose (DE-52) column chromatography, is activated 2-2.5-fold on incubation with 10 microM each of glucose-6-phosphate, NADPH, and glucose for 20 min at 25 degrees C. The activation of the enzyme was established by following the NADPH oxidation as well as the sorbitol formation using glucose as substrate. The activated form of aldose reductase exhibited monophasic kinetics with glucose and glyceraldehyde, whereas the unactivated or native enzyme exhibited a biphasic kinetics with both the substrates. The activated enzyme was less susceptible to inhibition by aldose reductase inhibitors such as sorbinil, alrestatin, and quercetrin as compared with the unactivated enzyme. Similarly, the native enzyme was strongly inhibited by some of the phosphorylated intermediates of glycolytic pathway, such as 3-phosphoglycerate, 1,3-diphosphoglycerate, 2,3-diphosphoglycerate, and ADP, whereas the activated enzyme was either not inhibited or inhibition was 20-30% only. Partially purified aldose reductase from the normal human lens exhibited properties similar to the native enzyme of other tissues, whereas the enzyme from clear lens obtained from diabetic subjects with severe hyperglycemia expressed properties similar to the in vitro activated enzyme of aorta, brain, and muscle.

Adult↗
PubMed1984

Aldose and aldehyde reductases in human tissues.

Immunochemical characterizations of aldose reductase and aldehyde reductases I and II, partially purified by DEAE-cellulose (DE-52) column chromatography from human tissues, were carried out by immunotitration, using antisera raised against the homogenous preparations of human and bovine lens aldose reductase and human placenta aldehyde reductase I and aldehyde reductase II. Anti-aldose antiserum cross-reacted with aldehyde reductase I, anti-aldehyde reductase I antiserum cross-reacted with aldose reductase and anti-aldehyde reductase II antiserum precipitated aldehyde reductase II, but did not cross-react with aldose reductase or aldehyde reductase I from all the tissues examined. DE-52 elution profiles, substrate specificity and immunochemical characterization indicate that aldose reductase is present in human aorta, brain, erythrocyte and muscle; aldehyde reductase I is present in human kidney, liver and placenta; and aldehyde reductase II is present in human brain, erythrocyte, kidney, liver, lung and placenta. Monospecific anti-alpha and anti-beta antisera were purified from placenta anti-aldehyde reductase I antiserum, using immunoaffinity techniques. Anti-alpha antiserum precipitated both aldehyde reductase I and aldose reductase, whereas anti-beta antibodies cross-reacted with only aldehyde reductase I. Based on these studies, a three gene loci model is proposed to explain the genetic interrelationships among these enzymes. Aldose reductase is a monomer of alpha subunits, aldehyde reductase I is a dimer of alpha and beta subunits and aldehyde reductase II is a monomer of delta subunits.

Aldehyde Oxidoreductases↗
PubMed1984

Interaction of silicate dust with erythrocyte ghost membrane: in vitro studies.

Composition of the erythrocyte ghost membrane obtained from slate-dust-lysed and hypotonically lysed erythrocytes were compared in vitro. The protein and cholesterol contents were unaltered, whilst phospholipid and glycosamine contents decreased significantly in the slate-dust-lysed preparation of erythrocyte ghost membrane. Na+, K+-ATPase activity remained unchanged, whilst acetylcholinesterase activity was decreased slightly by slate dust treatment. Further, binding of silicic acid, dissolving out of slate dust, was observed with a component of erythrocyte ghost membrane protein having molecular weight around 90 000 daltons. The significance of the findings is discussed.

Carrier Proteins↗
PubMed1984

The binding of silica to proteins from plasma and lungs of rat: in vitro.

Silica dissolving out from the slate dust was found to bind with plasma protein and purified bovine serum albumin. At 24 h of incubation at 37 degrees C binding affinity of silica (microgram of silica bound/mg of protein) with plasma protein and bovine serum albumin was found to be 0.59 and 0.44, respectively. By molecular exclusion chromatography using Sephadex G-200, silica binding protein of plasma was determined to be of mol. wt. around 67000. Similar proteins having silica binding capacity (mol. wt. 70000 and 85000) were also found in rat lung but these proteins unlike their plasma counterpart were glycoprotein in nature. Polyacrylamide gel electrophoresis of plasma and protein rich lung fraction show that proteins upon binding with silica undergo mobility changes. Significance of the existence of silica binding protein in plasma and lung of rat in relation to silica toxicity is discussed.

Animals↗
PubMed1983

Relationship between solubility and hemolytic effects of toxic dusts.

Two varieties of native and chemically treated slate dust were tested in vitro for their hemolytic effects and the extent of silicic acid dissolution in various physiological fluids. The extent of hemolysis was found to be proportional to the degree of dissolution of dust constituents. Membrane lysis by the dust appeared to be prevented by coating it with polyvinyl pyrrolidone, serum proteins and pulmonary lavage lipids. The significance of the findings is discussed.

Animals↗
PubMed1983

Lung mitochondria in experimental asbestosis.

Alterations in lung mitochondria were followed in guinea pigs at different periods after a single intratracheal injection of chrysotile dust. Cytochrome c oxidase and succinic dehydrogenase activities showed gradual increase after 90 days, whereas monoamine oxidase remained unaffected throughout the study. There was an increase in glutamate dehydrogenase activity in postmitochondrial as well as in mitochondrial fractions, the latter being accompanied by decreased latency of the enzyme. Mitochondria from asbestotic lung appeared to be more swollen than in normal animals at and after 90 days of exposure. There were fluctuations in the contents of different phospholipids as a result of asbestosis. Beyond 90 days, collagen and mucopolysaccharides also increased. The results confirm the contention that pulmonary mitochondria are among the major target sites in asbestosis.

Animals↗
PubMed1983

Biochemical studies on the toxicity of hematite dust.

Biochemical alterations in guinea pig lungs caused by hematite dust were followed at 150 days after intratracheal administration of the dust. In vivo dust exposure caused a significant increase in mitochondrial protein content and cytochrome c oxidase activity whereas diaphorase activity remained unaltered. Mitochondria from the exposed animals were apparently in a swollen state and their contraction profile upon the addition of ATP reflected permeability changes. However, in vitro dust caused no significant alterations. Significant increases in glycogen content along with an insignificant decrease in glycogen phosphorylase activity were also observed in hematite-treated guinea pig lungs. Decrease in drug-metabolizing enzymes such as aniline hydroxylase and tyrosine aminotransferase activities were also evident in the postmitochondrial fraction of the siderotic lungs. [3H]Leucine-incorporation studies showed increased protein synthesis in the postmitochondrial fraction. Increase in protein synthesis in mitochondria was only marginal whereas in whole homogenate it decreased considerably. Experiments employing dust tagged with radioactive iron indicated the rapid mobilization of iron from lung and its distribution to various organs. The presence of iron-binding protein was confirmed by employing Sephadex gel-filtration techniques.

Animals↗